There are three main objectives of the research program. First, we plan to carry out a detailed examination of the glutamine transport system of renal mitochondria. Using a rapid, Millipore-filtration assay system we will determine the kinetics of glutamine transport across the inner membrane of mitochondria-Km, Vmax, pH optimum, etc.-and develop an optimum assay medium to measure initial rates of glutamine uptake by renal mitochondria. We will use this method to determine the role of mitochondrial glutamine transport in the adaptation of renal ammonia excretion to metabolic acidosis. Second, we plan to determine whether skeletal muscle is a major site of glutamine synthesis in the rat and which biochemical pathway is used for the production of ammonia, a precursor of glutamine, in this tissue. Rates of glutamine synthesis by skeletal muscle will be determined in vitro and compared to the utilization of glutamine by the kidneys of normal and acidotic rats. The relative activities of the glutamate dehydrogenase pathway and the purine nucleotide cycle will be assayed under physiological conditions, in vitro, in order to determine which of the two pathways is more important in the production of ammonia in skeletal muscle. Third, we plan to determine the role of renal phosphate-dependent (PD) glutaminase in the maturation of the response of renal ammonia excretion to acidosis in infant rats. We will attempt to prevent the normal developmental rise in renal PD-glutaminase activity to deduce whether an increase in enzyme activity is a prerequisite for the maturation of the ammonia response. In a related study we will determine the role of PD-glutaminase in the response of ammonia excretion to acidosis in certain adult animals in which enzyme activity may be rate limiting in the production of urinary ammonia-uni-nephrectomized rats and herbivores (e.g. guinea pig). BIBLIOGRAPHIC REFERENCES: Goldstein, L., alpha-Ketoglutarate regulation of glutamine transport and deamidation by renal mitochondria. Biochem. Biophys. Res. Commun. 70: 1136-1141, 1976. Benyajati, S. and L. Goldstein. Glutaminase dependency of ammonia excretion adaptation in sub-total nephrectomized rats. Fed. Proc. 36: 605, 1977.